A well-appreciated need for labeling exists in society. Labeling is a fundamental basis for tracking and identifying. Encoding can be used as a form of labeling understood by persons or equipment, as in the case of bar coding. At the microscale, however, labeling/encoding itself becomes difficult.
Strategies to encode microscale materials have accordingly received increased attention for such uses as high-throughput screening in the fields of drug discovery, genetics screening, biomedical research, and biological and chemical sensing. Concurrent research strategies for measuring an increased number of analytes while minimizing the necessary sample volume have focused on either on-chip spatially differentiated arrays or encoded beads. Large arrays have been developed for biological and/or chemical sensing purposes by making use of positional encoding to register specific analyte responses. The main advantage of using an array over a conventional single analyte sensor is the ability to process and analyze a large number of analytes simultaneously. Positional arrays, however, can suffer from slow diffusion rates and limits on the concentration ranges of analytes being sensed. An alternative approach is to use individually encoded beads.
Early attempts to encode particles used fluorescent or infrared-active molecules as binary markers. More recently, cadmium selenide quantum dots have been demonstrated as viable candidates for particle encoding based on their unique fluorescent properties. Quantum dots have the advantage over organic molecules of increased stability towards photobleaching, sharper fluorescence peaks, improved solubility characteristics, and large excitation frequency ranges. With six colors (limited to the peak width of the fluorescence in the visible range) and ten intensity levels, 106 particles could theoretically be encoded. In practice, this number is difficult to obtain because of spectral overlap and sample inhomogeneities. Also, despite the increased photostability of quantum dots, fluorescence quenching is still possible, casting uncertainty on using relative intensity measurements as a reliable encoding method.
Another encoding strategy has used sub-micron metallic rods. The sub-micron metallic rods are prepared by electrodeposition of metals on a porous membrane in alternating strips of controlled thickness. Different reflection characteristics of the various metals are used as a barcode for identification purposes. Reflection spectroscopy does not have the disadvantage of photobleaching inherent with fluorophores. Additionally, fluorescent analytes do not interfere with the particle signal. Deposition of rods is a relatively complex process, however, and may be difficult to apply as an encoding strategy where, for example, a large number of codes is desirable because each rod must be brought into focus in an optical reader (such as a microscope) in order to read out the codes.
Fluorescent molecule encoding, core-shell quantum dot encoding, and photonic crystal encoding using Rugate and Bragg reflectivity theory rely upon creating spectral lines that act as bits. The number of possible codes is limited to 2n, where n is the number of spectral lines or bits that are discernable from other lines in a spectrum. There remains a need for encoding strategies at the microscale.